Studies in several animal models of human autoimmune diseases have revealed that with the progression of disease, there is a dynamic shift in the T cell responses to various epitopes within a particular self antigen (intramolecular) as well as to other disease-related self antigens (inter-molecular). This phenomenon has been termed as epitope spreading (or diversification of response), and it has been observed in experimental models [e.g., experimental autoimmune encephalomyelitis (EAE), type I diabetes in the non-obese diabetic (NOD) mouse, and systemic lupus erythematosus (SLE), etc.], and in patients with multiple sclerosis. In these disorders, epitope spreading has been invoked in propagation of the autoimmune response. In contrast to the above, using the rat adjuvant-induced arthritis (AA) model of human rheumatoid arthritis (RA), we observed that epitope spreading involving T cell response to 65-kD mycobacterial heat-shock protein (Bhsp65) occurred during the course of the disease, and that the T cell epitopes involved (namely, Bhsp65 C-terminal determinants; BCTD) were disease-regulating in nature. [AA is inducible in the Lewis rat by challenge s.c. with heat-killed M. tuberculosis (Mtb).] Thus, epitope spreading is not always involved in perpetuation of the autoimmune response; instead it can also be protective in nature. Defining the mechanisms underlying epitope spreading is critical for further understanding of the pathogenesis of autoimmunity, and for devising novel and more effective therapeutic approaches for these disorders. The AA model is most suited for study of epitope spreading in the setting of immune regulation. Nevertheless, information about the basic mechanisms involved in epitope spreading in AA would also be applicable to other models of autoimmunity. We hypothesize that diversification of the T cell response to BCTD is indeed triggered in vivo by self (rat) hsp65 (Rhsp65). It involves the enhanced cellular induction and expression of Rhsp65 coupled with upregulation of the antigen processing machinery under local inflammatory/cytokine milieu of acute AA, leading to efficient display of Rhsp65 C-terminal determinants (RCTD) from Rhsp65 and induction of RCTD-reactive T cells. These RCTD-reactive T cells are the ones that are recruited through crossreactivity (molecular mimicry) by homologous BCTD, manifesting as epitope spreading to Bhsp65 in the late phase of AA. We plan to address the following - Aim 1: To determine the role of self hsp65 (Rhsp65) and molecular mimicry in diversification of response (epitope spreading) to disease-regulating BCTD during the course of AA; and Aim 2: To study the influence on both clinical disease and epitope spreading of neonatal tolerization of the regulatory T cell repertoire, and to define the mechanism by which BCTD/RCTD-reactive T cells control the activity of arthritogenic epitope-specific T ceils. The results of this study would provide novel insights into the mechanisms underlying epitope spreading, and its role in regulation of autoimmunity. This in turn would contribute to developing novel therapeutic approaches for RA and other autoimmune diseases. [unreadable] [unreadable]